Antibody repertoire library

Eggena [17] used panning on neutrophil cells to isolate a specific clone from an antibody repertoire library prepared from an ulcerative colitis patient s lamina propria. This antibody represents an immunogenetic marker for the disease. 

The isolation of disease-related antibodies specific for either melanoma [ 180], the autoimmune thyroid peroxidase antigen [174], or neutrophil antigen characteristic of ulcerative colitis [17] from nonnaive patient repertoire phage libraries has been reviewed above in the Panning on cells section. The isolation of anti-viral antibodies with diagnostic or therapeutic potential from antibody repertoire libraries has been described in earlier papers e.g. against HIV [197], hepatitis B virus [198] and human respiratory syncytial virus [199],... 

Geoffroy, F., Sodoyer, R., Aujame, L (1994) A new phage display system to construct multicombinatonal libraries of very large antibody repertoires Gene 151, 109-113... 

In the past decade, the in vitro selection of antibodies from recombinant antibody repertoires has proven to bypass these limitations. In vivo libraries offer the capability of generating human antibodies in transgenic mice carrying the human IgG loci. Here, antigens are injected into the humanized mouse, followed by conventional hybridoma technology for selection and production of antibodies (4-6). 

Libraries of such size have so far only been generated for antibody repertoires. These are constructed by in vivo recombination of smaller, primary sublibraries generated by PCR (e.g., Sblattero and Bradbury, 2000). This level of sequence diversity may be attainable for other proteins, as long as they consist of multiple independent folding domains that can tolerate the insertion of recombination sites within the linker region. 

Figure 7.2 Profiling the antibody repertoire of autoimmune patients by combining protein microarray technology with large cDNA expression libraries. In a first step, serum samples from patients were analysed onto protein arrays consisting of approximately 37,200 redundant human proteins. The data sets obtained were compared to the data sets deriving from incubations with serum of age and sex matched control persons.

Researchers have expanded upon the natural diversity generated by the immune system. In 1989 a novel vector system based on the bacteriophage lambda was used to express a synthetic combinatorial library of Fab fragments of the mouse antibody repertoire in Escherichia coli [22]. This system allowed rapid and easy identification of monoclonal Fab fragments that bind a given antigen, in a form suitable for genetic manipulation. For example. 

Sastry, L., Alting-Mees, M., Huse, W.D., Short, J.M., Sorge, J.A., Hay, B.N., Janda, K.D., Benkovic, S.J., and Lerner, R.A. (1989) Cloning of the immunological repertoire in Escherichia coli for generation of monoclonal catalytic antibodies Construction of a heavy chain variable region-specific cDNA library. Proc. Natl. Acad. Sci. USA 86, 5728-5732. 

Methods of this nature are adequate for screening sets of hybridomas but not for selection from much larger libraries of antibodies. So, most recently, selection methods employing suicide substrates (Section 7) (Janda etal., 1997) or DNA amplification methodology (Fenniri et al., 1995) have been brought into the repertoire of techniques for the direct identification of antibodies that can turn over their substrate. However, the tedious screening of hybridomas remains the mainstay of abzyme identification. 

By similar logic, protein affinity libraries have been constructed to identify protein—protein combining sites, as in antibody—antigen interaction (19) and recombinant libraries have been made which produce a repertoire of antibodies in E. coli (20). In another case, a potential DNA-based therapeutic strategy has been studied (21). DNAs from a partially randomized library were sdected to bind thrombin in vitro. Oligonucleotides, termed aptamers that bound thrombin shared a conserved sequence 14—17 nudeotides long. 

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